Detection of matrix metallopeptidase-9-like proteins in Trypanosoma cruzi

作者：Nogueira de Melo Ana Cristina; de Souza Edilma Paraguai; Ramalho Elias Camila Guarany; Souza dos Santos Andre Luis; Branquinha Marta Helena; d'Avila Levy Claudia Masini; Garcia dos Reis Flavia Coelho; Rocha Costa Tatiana Ferreira; Cabral de Araujo Lima Ana Paula; de Souza Pereira Mirian Claudia; Leal Meirelles Maria Nazareth; Vermelho Alane Beatriz^*

来源：Experimental Parasitology, 2010, 125(3): 256-263.

DOI：10.1016/j.exppara.2010.01.023

• Trypanosoma cruzi
• Peptidase
• Matrix metallopeptidase-9
• Doxycycline

In this study, the cell-associated and extracellular peptidases of Trypanosoma cruzi grown in modified Roitman's complex (MRC) medium were analyzed by measuring peptidase activity in gelatin-containing zymograms. Our results showed that the cell-associated peptidases as well as peptidases extracellularly released by T. cruzi displayed two distinct proteolytic classes: cysteine and metallopeptidase activities. The major cysteine peptidase, cruzipain, synthesized by T. cruzi cells was detected in cellular parasite content, as a 50 kDa reactive polypeptide, after probing with anti-cruzipain antibody. In addition, metallo-type peptidases belonging to the matrix metallopeptidase-9 (MMP-9) family were revealed, after Western blotting, as a 97 kDa protein band in cellular extract and an 85 kDa polypeptide in both cellular and secreted parasite extracts. The MMP-9-like activity present in cells and spent culture medium was immunoprecipitated by an anti-MMP-9 polyclonal antibody. The surface location of MMP-9-like proteins in T. cruzi was also evidenced by means of flow cytometry analysis. Furthermore, doxycycline that has direct MMP-9 inhibiting properties in vitro, inhibited MMP-9-like activities in gel zymography, immunoprecipitation and flow cytometry analyses. This is the first report of the presence of MMP-9-like molecules in T. cruzi. The presence of a matrix extra\cellular-degrading enzyme may play a role in the T. cruzi-host cell interaction, making this enzyme a potential target for future drug development against this pathogenic trypanosomatid.

• 出版日期2010-7